The cells of the intestinal epithelium are an active partner in mucosal immunity and comprise a self renewing physical barrier that is essential to minimizing exposure to foreign and toxic stimuli from the external environment. Chemokines are ubiquitous regulatory factors that participate in the activation and directional trafficking of leukocytes, and are a significant component of the physiologic mucosal immune response. The biologic function of these molecules is mediated through chemokine receptors, which stimulate migration, proliferation and differentiation in specific target cells. Intestinal epithelial cells express a monogamous chemokine/chemokine receptor pair that has been shown to be essential in vivo. The overall objective of this proposal is to explore the role of the chemokine receptor CXCR4 in establishing innate mucosal host defense though the regulation of epithelial barrier integrity. To date, investigations into the physiologic role of CXCR4 in the gastrointestinal mucosa have been prevented by the embryonic lethality of CXCR4 gene deficient mice. We will use both tissue culture models of intestinal epithelia and conditional knockout mice to mechanistically define CXCR4 management of the mucosal barrier. Studies in Aim 1 will employ human and rat intestinal epithelial culture models to test the hypothesis that CXCR4 signals via coupled G-proteins and PI3K to regulate epithelial restitution and establish a secure epithelial barrier. In Aim 2, we will extend our studies by testing the hypothesis that CXCR4 regulates barrier maintenance indirectly; through G-protein blockade of cAMP, and directly, through Rho-activated modulation of the actin cytoskeleton and cell-cell junctions. In Aim 3 we will use conditional knockout mice to test the hypothesis that CXCR4 is a critical effector of intestinal epithelial migration and barrier morphogenesis in vivo. These studies will be the first to detail functions for the chemokine receptor CXCR4 as a regulator of mucosal barrier homeostasis and wound repair. Elucidation of the biochemical and cellular mechanisms regulating this process will have great relevance to the design of therapeutic strategies to manage intestinal damage sustained as a result of infectious disease or chronic inflammatory disorders.